The present invention relates to isolated polynucleotide molecules useful for analyzing cardiac potassium channel minK subunit variants, and to screening and diagnostic uses thereof relating to a polymorphism in the KCNE1 gene encoding the cardiac potassium channel minK subunit polypeptide. Among such uses are methods for determining the susceptibility of a subject to drug-induced cardiac arrhythmias based on an analysis of a biological sample isolated from the subject.
Cardiac arrhythmias are a cause of substantial morbidity and mortality in adults. A variety of therapeutic agents commonly used to treat arrhythmias, along with other non-cardiac drugs, sometimes provoke potentially dangerous disturbances of cardiac rhythm. Prolongation of the cardiac action potential by blocking the rapidly activating cardiac delayed rectifier potassium current, IKr, is the desired therapeutic effect of many anti-arrhythmic agents, but is an inadvertent adverse effect of certain antihistamines, antidepressants, gastric motility promoters, and other agents. Generally without warning, 1-10% of patients receiving action potential prolonging drugs will develop marked prolongation of the electrocardiographic QT interval or polymorphic ventricular tachycardia (torsades de pointes). See Roden, D. M., N.Engl.J.Med. 331:785-791 (1994); Roden, D. M., Am J Cardiol 72:44B-49B (1993); Carlsson, L., et al., J. Pharmacol. Exp. Ther. 282:220-227 (1997); Mohammad, S., et al., Am. J. Physiol. Heart Circ. Physiol. 273:H2534-H2538(1997); Rampe, D., et al., FEBS Lett 417:28-32 (1997); Woosley, R. L., et al., JAMA 269:1532-1536 (1993); Suessbrich, H., et al., FEBS Lett 385:77-80 (1996); Weissenburger, J., et al., Clin Exp Allergy 29 (Suppl. 3):190-196 (1999); Jackman, W. M., et al., Prog Cardiovasc Dis 31:115-172 (1988); Lazzara, R. Eur Heart J 14:H88-H92(1993); Tan, H. L., et al., Ann.Intern.Med. 122:701-714 (1995).
Congenital long QT syndrome (LQTS) is an inherited condition of abnormal cardiac repolarization characterized clinically by an increased risk of torsades de pointes. See Keating, M. T. Medicine (Baltimore) 75:1-5 (1996); Vincent, G. M. Annu. Rev. Med. 49:263-274 (1998); Roden, D. M., et al., Circulation 94:1996-2012 (1996). The majority of LQTS subjects appear to harbor mutations in either of two cardiac potassium channel genes, HERG and KCNQ1 (see Curran, M. E., et al., Cell 80:795-803 (1995) and Wang, Q., et al., Nature Genet 12:17-23 (1996)), while additional cases are caused by mutations in genes encoding potassium channel regulatory subunits KCNE1 and KCNE2 (see Splawski, I., et al., Nature Genet. 17:338-340 (1997); Abbott, G. W., et al., Cell 97:175-187 (1999)), a cardiac voltage-dependent sodium channel xcex1-subunit SCN5A (see Wang, Q., et al., Cell 80:805-811 (1995)), and other unidentified gene products (see Schott, J. J., et al., 57:1114-1122 (1995)).
A few anecdotal reports have attempted to associate the presence of rare sequence variants in KCNQ1 and KCNE2 to drug-induced LQT in the absence of an overt congenital phenotype. See Abbott, G. W., et al., Cell 97:175-187 (1999); Schulze-Bahr, E., et al., Circulation 96:1-211(1997); Napolitano, C., et al., Circulation 96:1-211(1997); Donger, C., et al., Circulation 96:2778-2781 (1997); Priori, S. G., et al., Eur Heart J 18:324(1997). However, in these cases, the described alleles were absent in the general population, indicating that they are only rare causes of drug-induced arrhythmia susceptibility. Thus, the prevalence of drug induced cardiac arrhythmias in the general population remains unexplained and uncharacterized.
Therefore, drug-induced cardiac arrhythmias continue to represent the Achilles heel of efforts to develop safe and effective anti-arrhythmic agents. Moreover, drug induced cardiac arrhythmias also occur during treatment with non-cardiac drugs that have unintended effects on cardiac repolarization. For example, as many as 10% of patients treated with quinidine, sotalol, and ibutilide will develop excessive QT interval prolongation or exhibit the precipitous occurrence of torsade de pointes. This unpredictable adverse reaction can occur in the absence of identifiable risks factors such as hypokalemia, hypomagnesemia, concomitant treatment with other IKr blockers, and recent conversion from atrial fibrillation. See Tan, H. L., et al., Ann. Intern. Med. 122:701-714 (1995).
A method that can predict individual susceptibility to drug-induced arrhythmias would have substantial clinical utility and would meet a long-felt need in the art. However, such a method is currently not available in the art.
A method of screening for susceptibility to a drug-induced cardiac arrhythmia in a subject is disclosed. The method comprises: (a) obtaining a biological sample from the subject; and (b) detecting a D85N polymorphism of a KCNE1 gene encoding a cardiac potassium channel minK subunit polypeptide in the biological sample from the subject, the presence of the D85N polymorphism indicating that the susceptibility of the subject to a drug-induced cardiac arrhythmia.
Preferably, the polymorphism of the minK polypeptide comprises a G to A transition in the single exon of the KCNE1 gene, more preferably at nucleotide 281 of a cDNA that corresponds to the KCNE1 gene. More preferably, the G to A transition at nucleotide 281 of the cDNA that corresponds to the KCNE1 gene further comprises a change in the triplet code from GAC to AAC or GAT to AAT, which encodes a KCNE1 polypeptide having an asparagine (Asn or N) moiety at amino acid 85, instead of an aspartate (Asp or D) moiety. Hence, the polymorphism is referred to as the KCNE1-D85N polymorphism.
Kits and reagents, including oligonucleotides, nucleic acid probes and antibodies suitable for use in carrying out the methods of the present invention and for use in detecting minK polypeptides and KCNE1 polynucleotides are also disclosed herein.
It is therefore an object of the present invention to provide polynucleotide molecules that can be used in analyzing a cardiac potassium channel minK subunit gene (KCNE1) in vertebrate subjects.
It is also an object of the present invention to provide for the determination of KCNE1 genotype in vertebrate subjects and particularly human subjects, based on information obtained through the analysis of nucleic acids, including genomic DNA and cDNA, derived from tissues from the subject.
It is yet another object of the present invention to provide a ready method for determining KCNE1 genotype.
It is still a further object of the present invention to provide polypeptide and polynucleotide molecules for use in generating antibodies that distinguish between the different forms of KCNE1 which constitute the KCNE1-D85N polymorphism.
It is yet a further object of the present invention to provide methods for diagnosing clinical syndromes related to and associated with the KCNE1-D85N polymorphism.
Some of the objects of the invention having been stated hereinabove, other objects will become evident as the description proceeds, when taken in connection with the accompanying Drawings and Examples as best described hereinbelow.